Large-scale serological survey on Mycobacterium avium subsp. paratuberculosis infection in sheep and goat herds in Sicily, Southern Italy.

Introduction: Paratuberculosis (PTB) is a worldwide chronic, contagious enteric disease caused by Mycobacterium avium subsp. paratuberculosis (MAP) mainly affecting ruminant species. PTB is a WOAH-listed disease with direct and indirect economic losses in the livestock sector, negative impact on animal welfare and significant public health concerns. In spite of this, MAP prevalence in small ruminants is still unknown and the prevalence appears to be underestimated in many countries. The aim of this study is providing a first large-scale serological survey on MAP infection in small ruminants in Sicily, a region of Southern Italy with the 11.3 and 8.9% Italian national heritage of sheep and goats, respectively. Methods: For this purpose, we analyzed a total of 48,643 animals reared in 439 flocks throughout Sicily. MAP seroprevalence was estimated both at herd-level and animal-level within breeds reared in all the nine sampled provinces. Results: Our results revealed a high overall apparent prevalence at herd-level of 71.8% in sheep and 60.8% in goat farms with an animal-level prevalence of 4.5 and 5.1% in sheep and goats, respectively. Significant statistical differences were found between the provinces and within the breeds both in sheep and goats. Discussion: Our study provides the first large-scale serological survey on PTB infection in small ruminants in Sicily and showed a high prevalence of disease depending to the species, breed and province. This study represents the first step to better understand the MAP epidemiology in a typical Mediterranean breeding context, suggesting the need of in-depth study on the herds risk factors, including the eventual presence of candidate genes for resistance/susceptibility to PTB in native breeds.

Infection Due to Actinobacillus lignieresii in Cattle with Brain and Ocular Involvement: Histological and Microbiological Features.

The actinobacillosis is rare and to date the biological profile of the agent is not yet fully understood. The knowledge about the possible hosts of the pathogen is incomplete and is generally only associated with granulomatous lesions in cattle and sheep. The primary organs involved are the mouth, tongue and pharynx. Human infection is extremely rare. Actinobacillus lignieresii is the causative agent of a rare bovine granulomatous disease known as "wooden tongue". In this research, we describe a case of cerebral and ocular metastatic diffusion of granuloma due to infection with Actinobacillus lignieresii in cattle, probably resulting from primary oral localization. Diagnosis was made using histopathological assay which made it possible to highlight the typical lesion of actinobacillosis, and bacteriological assay that allowed to isolate the pathogen.

Assessment of selected molecular factors and 17-ß estradiol dosage in response to Toxoplasma gondii infection in swine.

Background and Aim: Toxoplasma gondii is a global zoonotic parasite infecting virtually all warm-blooded species, although a species-specific variability is evident referring to symptoms frame. Both the success of T. gondii and the outcome of infection depend on a delicate balance between host cellular pathways and the evasion or modulation strategies elicited by the parasite. The hormonal and molecular mechanisms involved in this delicate host-parasite balance are still unclear, especially when considering intermediate host species other than mouse. This study aimed to assess any correlation between T. gondii infection and selected molecular and hormonal factors involved in responses to infection in susceptible species such as swine. Moreover, blood counts and hematochemical assays (glucose, total cholesterol, and triglycerides dosage) were performed to evaluate the overall health condition of animals. Materials and Methods: Toxoplasmosis was diagnosed by enzyme-linked immunosorbent assay for antibodies determination and real-time polymerase chain reaction (RT-PCR) for T. gondii DNA detection. Target genes coding for key factors of cell responses to T. gondii infection were selected, and their transcription was assessed in various tissues by quantitative RT-PCR. 17-ß estradiol concentrations were assessed by fluorimetric enzyme-linked immunoassay and the AIA-360 automated immunoassay analyzer. Blood count and hematochemical analyses were performed by a blood cell counter and a spectrophotometer, respectively. Results: The present research highlighted significant differences among infected and uninfected swine (control group) for both transcription profiles of some of the molecular factors considered and 17-ß estradiol concentrations. Referring to the assessed hematological and biochemical parameters, no statistically significant differences were observed in infected swine compared to the control group. Conclusion: Our results contribute to the enrichment of data available about the subject and could be useful for a deeper knowledge of the interaction between this parasite and its hosts. However, more aspects are still unclear, such as the effective response of downstream molecules from the same pathways to the variation of factors observed in this study either assessing how the same factors respond to Toxoplasma gondii infection in other host speciesand further analyses should be performed on other host species.

Edible Insects and Toxoplasma gondii: Is It Something We Need To Be Concerned About?

ABSTRACT: Novel foods, such as edible insects and food products on the basis of insects, could play an important role in both human and animal nutrition in the future. The identification of dangers associated with insect consumption is fundamental to guarantee consumer safety and adequate regulatory guidelines for operators of the food sector. Although former studies have focused on the microbiological contamination of fresh or processed edible insects, so far little information is available about the occurrence of foodborne parasites, such as Toxoplasma gondii, whose life cycles make them candidates for potential insect breeding substrate contamination. Hence, we investigated the presence of contaminating T. gondii in farmed edible insects to rule out this further hazard for consumers. Four species of insects most commonly used as food for human consumption were analyzed: mealworm; African migratory locust, house cricket, and silkworm. Samples included live specimens but also minimally (dehydrated) and highly processed edible insects. Traces of T. gondii DNA were detected in samples of dehydrated mealworm. These results highlight the need for implementing good farming and processing practices with particular care paid to safe storage and handling of feed and substrates used for edible insects to reduce the chance of T. gondii entering the human food chain.

Toxoplasma gondii seroprevalence in beef cattle raised in Italy: a multicenter study.

Toxoplasmosis represents an important public health issue, with the consumption of raw or undercooked meat being a major way of human infection. The role of beef in the transmission of the parasite to humans is questioned due to lower quantity of tissue cysts compared with other meat-producing species. However, the habit of consuming raw beef is regionally diffused, and the risk posed by Toxoplasma gondii infection in cattle should not be overlooked. Therefore, to update information on T. gondii in cattle reared in Italy, a multicentric seroepidemiological survey was designed and implemented in four Northern regions (Liguria, Lombardy, Piedmont, and Trentino Alto Adige) and Sardinia. Overall, a convenience sampling was performed, collecting 1444 serum samples from 57 beef cattle herds. Thirteen beef breeds were sampled, besides cross-breed; bovines age varied from 3 months to over 12 years. Sera were tested with a commercial ELISA for the detection of anti-T. gondii antibodies. Individual and herd data were analyzed by binary logistic regression analysis. A T. gondii seroprevalence of 10.2% was recorded, with differences among regions and values ranging from 5.3% in Liguria to 18.6% in the Piedmont region (p value = 0.0001). Both young and adult animals and males and females tested positive, without any significant difference (age and gender: p value > 0.05). Lower seroprevalence values were recorded in cattle born in Italy (8.7%) if compared with animals imported from abroad (13.4%) (p value = 0.046). The spread of T. gondii in beef cattle destined to Italian consumers is confirmed, suggesting the need of continuous monitoring of the infection.

Toxoplasma gondii in edible fishes captured in the Mediterranean basin.

The issue of whether market fish can be involved in the transmission of Toxoplasma gondii in the marine environment is highly debated since toxoplasmosis has been diagnosed frequently in cetaceans stranded along the Mediterranean coastlines in recent times. To support the hypothesis that fishes can harbour and effectively transmit the parasite to top-of-the-food-chain marine organisms and to human consumers of fishery products, a total of 1,293 fishes from 17 species obtained from wholesale and local fish markets were examined for T. gondii DNA. Real-time PCR was performed in samples obtained by separately pooling intestines, gills and skin/muscles collected from each fish species. Thirty-two out of 147 pooled samples from 12 different fish species were found contaminated with T. gondii DNA that was detected in 16 samples of skin/muscle and in 11 samples of both intestine and gills. Quantitative analysis of amplified DNA performed by both real-time PCR and digital PCR (dPCR) confirmed that positive fish samples were contaminated with Toxoplasma genomic DNA to an extent of 6.10 × 10-2 to 2.77 × 104  copies/ml (quantitative PCR) and of 1 to 5.7 × 104  copies/ml (dPCR). Fishes are not considered competent biological hosts for T. gondii; nonetheless, they can be contaminated with T. gondii oocysts flowing via freshwater run-offs (untreated sewage discharges, soil flooding) into the marine environment, thus acting as mechanical carriers. Although the detection of viable and infective T. gondii oocysts was not the objective of this investigation, the results here reported suggest that fish species sold for human consumption can be accidentally involved in the transmission route of the parasite in the marine environment and that the risk of foodborne transmission of toxoplasmosis to fish consumers should be further investigated.

Evaluation of a commercial enzyme-linked immunosorbent assay (ELISA) for detecting antibodies against Toxoplasma gondii from naturally and experimentally infected pigs.

BACKGROUND: Toxoplasmosis is one of the most frequent parasitic infections in animals causing reproductive disorders and thus notable economic losses in productivity. Among food animals, pigs along with sheep and goats possess the highest incidence of Toxoplasma gondii cysts in meat, and play a role as a source of human infection. METHODS: The commercial ELISA kit (PrioCHECK® Toxoplasma Ab SR, Prionics Schlieren-Zurich, Switzerland) for the detection of specific antibodies against T. gondii in swine serum was compared with a commercial IFAT (indirect fluorescent antibody test) (Toxo-Spot IF, bioMérieux, France), used as the reference test. RESULTS: The kappa value obtained comparing the results performed on sera by ELISA with the by IFAT was 1. By a receiver operating characteristics curve analysis, the commercial ELISA had a relative sensitivity of 100%, and a relative specificity of 100% respect to IFAT. CONCLUSIONS: The commercial ELISA showed a very good agreement with the commercial IFAT in the detection of serum antibodies to Toxoplasma in pigs. Our study confirmed the usefulness of the commercial ELISA kit for the detection of antibodies against T. gondii in pigs, representing a valuable tool to improve the diagnostic activity for T. gondii in swine populations at the farm level or at the slaughterhouse, contributing to the control of this widespread infection.

The effect of Toxoplasma gondii on plasma serotonin concentration in sheep.

BACKGROUND AND AIM: Toxoplasma gondii is an intracellular parasite that commonly infects warm-blooded animals, including humans. Virtually all species can be infected, but a species-specific variability is evident, in terms of both type and severity of the symptoms encountered. As serotonin (5-hydroxytryptamine [5-HT]) plays an important regulatory role in both physiological and immune responses, the aim of this research was to assess whether toxoplasmosis disease could affect plasma 5-HT concentration and/or hematochemical parameters in a particularly susceptible species to infection as sheep. MATERIALS AND METHODS: 5-HT plasma levels were analyzed in platelet-poor plasma fraction by enzyme-linked immunosorbent assay. Blood count and hematochemical parameters were evaluated. Total proteins (TPs), glucose (Glu), and lactate dehydrogenase were determined by a spectrophotometer. RESULTS: Results showed significantly higher levels in plasma 5-HT, monocytes, and TP and significantly lower levels of Glu, in infected sheep compared to the control group. CONCLUSION: Results could support the hypothesis of an effect of toxoplasmosis infection on plasma 5-HT concentrations in sheep. More research is needed to assess the function of 5-HT in the regulation of infected sheep's immune responses.

Development and validation of a real-time PCR assay for the detection of Toxoplasma gondii DNA in animal and meat samples.

We report a rapid and reliable method for the detection of Toxoplasma gondii in meat and animal tissues based on real-time polymerase chain reaction (PCR). Samples were collected from cattle, small ruminants, horses, and pigs raised or imported into Sicily, Italy. All DNA preparations were assayed by real-time PCR tests targeted to a 98-bp long fragment in the AF 529-bp repeat element and to the B1 gene using specific primers. Diagnostic sensitivity (100%), diagnostic specificity (100%), limit of detection (0.01 pg), efficiency (92-109%), and precision (mean coefficient of variation = 0.60%), repeatability (100%), reproducibility (100%), and robustness were evaluated using 240 DNA extracted samples (120 positives and 120 negative as per the OIE nested PCR method) from different matrices. Positive results were confirmed by the repetition of both real-time and nested PCR assays. Our study demonstrates the viability of a reliable, rapid, and specific real-time PCR on a large scale to monitor contamination with Toxoplasma cysts in meat and animal specimens. This validated method can be used for postmortem detection in domestic and wild animals and for food safety purposes.

A clinical case of dourine in an outbreak in Italy.

In May 2011, dourine was reported in Italy following the declaration of a positive result observed in a stallion undergoing routine testing for stud purposes. Clinical signs, anatomo-histopathological findings and laboratory results that resulted in the confirmation of diagnosis of dourine in a clinically affected mare, which was the likely source of infection in the stallion, are described.